Hayashi S, Chang S Y, Chang S, Giam C Z, Wu H C
J Biol Chem. 1985 May 10;260(9):5753-9.
We have cloned the Escherichia coli lipoprotein structural gene (lpp) into a shuttle vector and studied its expression in both E. coli and in Bacillus subtilis. Using in vitro gene fusion techniques, the lpp gene was placed under the control of the promoter for the erythromycin-resistance (ery) gene. This fusion gene directed the synthesis of Braun's prolipoprotein which can be subsequently processed into the mature lipoprotein. In addition to the prolipoprotein, two ery-lpp hybrid proteins containing a 45- and a 22-amino acid extension preceding the NH2 terminus of prolipoprotein, respectively, are also synthesized in E. coli. The synthesis of these three proteins appears to involve the utilization of three distinct translation initiation sites. In B. subtilis, only two proteins are synthesized, the hybrid protein with a 45-amino acid extension and the prolipoprotein. In both E. coli and B. subtilis, the precursor forms of the hybrid proteins are lipid-modified, and they are processed to mature lipoprotein in vivo. These results indicate that internalized signal sequence containing the prolipoprotein modification and processing site (Leu-Ala-Glys-Cys) can function normally and permit the modification of hybrid proteins to lipid-modified precursors which can be subsequently processed by the globomycin-sensitive prolipoprotein signal peptidase.
我们已将大肠杆菌脂蛋白结构基因（lpp）克隆到一个穿梭载体中，并研究了其在大肠杆菌和枯草芽孢杆菌中的表达情况。利用体外基因融合技术，将lpp基因置于红霉素抗性（ery）基因启动子的控制之下。这个融合基因指导合成布劳恩前脂蛋白，随后可将其加工成成熟脂蛋白。除了前脂蛋白外，在大肠杆菌中还合成了两种ery-lpp杂合蛋白，它们在脂蛋白前体的NH2末端之前分别含有45个和22个氨基酸的延伸序列。这三种蛋白质的合成似乎涉及利用三个不同的翻译起始位点。在枯草芽孢杆菌中，只合成了两种蛋白质，即具有45个氨基酸延伸序列的杂合蛋白和前脂蛋白。在大肠杆菌和枯草芽孢杆菌中，杂合蛋白的前体形式都进行了脂质修饰，并在体内加工成成熟脂蛋白。这些结果表明，包含前脂蛋白修饰和加工位点（亮氨酸-丙氨酸-甘氨酸-半胱氨酸）的内化信号序列能够正常发挥功能，并允许将杂合蛋白修饰为脂质修饰的前体，随后可由对球霉素敏感的前脂蛋白信号肽酶进行加工。
